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Environmental fate & pathways

Hydrolysis

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Reference
Endpoint:
hydrolysis
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 111 (Hydrolysis as a Function of pH)
Deviations:
no
GLP compliance:
no
Analytical monitoring:
yes
Details on sampling:
- Sampling intervals for the parent/transformation products:
pH 1.2: The recordings were done at t = 3 min; 7 min; 39 min; 142 min; 442 min; 1293 min.
pH 4: The recordings were done at t = 3 min; 37 min; 97 min; 137 min; 446 min; 1329 min.
pH 7: The recordings were done at t = 3 min; 20 min; 51 min; 72 min; 372 min; 672 min.
pH 9: The recordings were done at t = 3 min; 10 min; 18 min; 44 min; 144 min; 287 min; 986 min

Due to the complexity of the overall hydrolysis process it was hard to determine the reaction time necessary for the best description. The reactions were followed for approximately 24 hours. It proved to be a good compromise, because in terms of the transformation of the test item the process was complete, but the transformation of the reaction products was still going on.

- Sampling method: The samples were kept isolated from light an oxygen in a drying oven at 25 ± 0.5°C (except at pH 1.2 which will be kept at 37°C). 1H-NMR measurements were run consecutively using the same duplicate samples on a Bruker NMR spectrometer equipped with a z-gradient probehead.
Buffers:
- pH: 1.2 (gastric), 4, 7 and 9
- Type and final molarity of buffer:
pH 1.2: The gastric pH was set with deuterochloric acid (DCl)
pH 4: 0.40 ml. 0.1 N NaOH +50 ml. biphthalate diluted to 100 ml
pH 7: 29.63 ml. 0.1 N NaOH + 50 ml. phosphate diluted to 100 ml
pH 9: 21.30 ml. 0.1 N NaOH + 50 ml. boric acid diluted to 100 ml
- Composition of buffer:
The concentrations of the buffers were 0.02 M. No additional organic solvent was added to the solutions.

Estimation method (if used):
For the kinetic calculations the following considerations were taken. The processes – even the complicated reactions – can be approximated with consecutive first order reactions where: [VL3]= VL30 e-kt
Where:
[VL3]: actual concentration of VL3,
VL30: initial concentration of VL3,
k: first order reaction constant
t: reaction time

The logarithmic form of this equation says: ln [VL3] = ln A0 + kt
And since the signal intensity is proportional to the concentration of the test item: ln [IVL3] = ln I0VL3 + kt
This is generally applicable for the cases where the decrease of the VL3 signal can be detected for example the process at pH=1.2, 7.0.

In the cases where the formation of the ethyl lactate was followed (pH=4.0,7.0 and 9.0) we mostly found complicated kinetic behaviour. In these cases two step reactions took place: a first fast transformation (process 1, like an almost immediate hydrolysis of the first ethyl lactate arm) and a second much slower (process 2, the deliberation of the second and possibly the third ethyl lactate arms). In these cases biexponetil fitting was applied and the following equation was applied for the determination of the ‘k’ values: [A]= A∞ -A1*exp(-k1*t)-A2*exp(-k2*t)
Where
[A]: actual concentration of hydrolysis product A
A∞: concentration of hydrolysis product A at the end of the reaction
A1 and A2: the fractions of hydrolysis product A participated in process 1 and 2
Details on test conditions:
TEST SYSTEM
The 0.1 mol/l NaOH and all other solutions and the final buffers were prepared in 100 ml “A” signed volumetric flasks made of glass. The dissolution of the chemicals was aided by 5 minutes treatment in ultrasonic bath. The sample solutions were made directly before spectrum recording in the 5 ml NMR tubes. The VL was weighted into the tubes with 0.1 mg precision.

TEST MEDIUM
Approximately 0.01 M solutions of the test item were prepared in buffers deoxygenated (by bubbling nitrogen through the buffers).
Duration:
24 h
pH:
1.2
Temp.:
37 °C
Initial conc. measured:
0.01 mol/L
Duration:
24 h
pH:
4
Temp.:
25 °C
Initial conc. measured:
0.01 mol/L
Duration:
24 h
pH:
7
Temp.:
25 °C
Initial conc. measured:
0.01 mol/L
Duration:
24 h
pH:
9
Temp.:
25 °C
Initial conc. measured:
0.01 mol/L
Positive controls:
no
Negative controls:
no
Preliminary study:
Tier 1: At the start of the reaction an almost immediate hydrolysis step takes place, where polymeric species form. This is indicated by the appearance of wide, unresolved signals. None of the solutions examinated proved to the stable over 5 days at 25 ºC.
Transformation products:
yes
No.:
#1
No.:
#2
No.:
#3
No.:
#4
Details on hydrolysis and appearance of transformation product(s):
- Formation and decline of each transformation product during test: Yes.
- Pathways for transformation: See below for details.
Key result
pH:
1.2
Temp.:
37 °C
Hydrolysis rate constant:
0.2 min-1
DT50:
3.46 min
Key result
pH:
4
Temp.:
25 °C
Hydrolysis rate constant:
0.15 min-1
DT50:
4.62 min
Key result
pH:
7
Temp.:
25 °C
Hydrolysis rate constant:
0.035 min-1
DT50:
19.8 min
Key result
pH:
9
Temp.:
25 °C
Hydrolysis rate constant:
0.073 min-1
DT50:
9.5 min
Details on results:
The test item hydrolyses very fast at pH 1.2 (37 ºC), 4 (25 ºC), 7 (25 ºC) and 9 (25 ºC) being the half-lives 3.46, 4.62, 19.8 and 9.5 min respectively.

MAJOR TRANSFORMATION PRODUCTS
At pH 1.2:
At the beginning (3 min), the signals of the NMR trace are not well resolved. This is due to the presence of other molecules (formed hydrolysis products: oligomers and polymers) in the size range where the molecules still provide NMR signal, but the peak widening effect of the colloidal particles is very much pronounced. As the reaction evolves the signals become resolved. From 7 minutes the spectrum “cleans up” and in the in the fourth spectrum (442 min), the main molecules present become visible. The first step is the mixing and the reaction with water. In acidic medium the hydrolysis is fast. The first NMR spectrum was recorded after 3 minutes of mixing. In this period already fast changes take place even during the scanning. The next spectrum at 7 minutes shows that the ethyl-lactate has been released mostly. According to the spectra in the first seven minutes of the reaction 2.4 mg ethyl-lactate was formed calculated from the CH3 methyl peak and 2.5 mg calculated from the CH2 peak. Since 3.5 mg VL3 was weighted into the sample which should result in 3.1 mg ethyl-lactate it can be stated that 75 % of the ethyl lactate was released from the VL3 in the first 7 minutes. Assuming that this process is first order it is possible to estimate a rate constant as follows: The decrease of VL3 in time from c0 concentration to its 25 % in 7 minutes is c0×0.25=c0exp(-k*7), being k = 0.2 min-1. The formation of ethyl-lactate has the same rate constants. After this fast reaction step the slow hydrolysis of ethyl-lactate to lactic acid and ethanol occurs with rate constant from parameter fitting 3.4 × 10-3 min-1. Since VL3 is almost completely hydrolysed in the beginning of the spectrum recording only amount of the hydrolysis product can be quantitated. The concentrations at each time points are summarized in Table 1 while the actual amounts expressed as % (n/n) and % (m/m) of the original VL3 amount in Table 2 and 3.

At pH 4:
As at pH 1.2 the bottom spectrum (t =3 min) differs in resolutions from the rest. The reason is the same and it was observed that after addition of VL3 slight opalescence appeared. After the addition of the test item into the buffer, VL3 releases the ethyl-lactate and starts to condensate to vinyl-ethyl-lactate colloid resulting in a decrease of VL3 concentration. Then the insoluble VL3 product undergoes further hydrolysis resulting in partial dissolution back to the solution but in the same time it starts to hydrolyse and release further free ethyl lactate to the solution. Approximately 2.5 mg (ca. 50 %) of the initial 4.7 mg VL3 precipitates in the first fast step and then colloidal VL3 slowly releases the ethyl-lactate. The kinetics can also be followed by following the formation of free ethyl-lactate. The two steps reaction is not as clearly visible as in the case of the decreasing peaks of coordinated ethyl-lactate. 0.4 mg (10 %) of the theoretical amount of maximum 4 mg ethyl-lactate is formed while the 4 mg ethyl-lactate forms in approximately 1400 minutes. At this pH VL3 hydrolysed only to ethyl lactate. No ethanol or lactic acid was found in quantity higher than 1%. The concentrations at each time points are summarized in Table 4 while the actual amounts expressed as % (n/n) and % (m/m) of the original VL3 amount in Table 5 and 6.

At pH 7:
In contrast to the acidic solutions the bottom spectrum (t =3 min) does not differ in resolutions from the later ones, and much less opalescence was observed at the sample preparation. The probable mechanism is that the first hydrolysis step is fast and results in consecutive loss of the ethyl lactate arms from VL3 and in formation of oligomers. It is supported by the same rate of decrease of VL3 and the increase of ethyl lactate. The rest of ethyl- lactate probably dissolves out of the poly-vinyl-siloxan in a slower step during the condensation. The trends can be described with two exponential functions providing an average rate constant of 0.04 min-1 is obtained and fora slower one 3 × 10-4 min-1. At this pH VL3 hydrolysed only to ethyl lactate. No ethanol or lactic acid was found in quantity higher than 1%. The concentrations at each time points are summarized in Table 7 while the actual amounts expressed as % (n/n) and % (m/m) of the original VL3 amount in Table 8 and 9.

At pH 9:
The higher pH has a positive effect on the dissolution of the test item hence on the quality of the spectra. This is supported by the fact that the initial spectrum at the bottom of the figure (t =3 min) does not differ in resolutions from the rest, and that after addition of VL3 a slight opalescence appeared which disappeared quickly. The hydrolysis of VL3 in alcalic medium takes place in three steps. First quickly, within 40 minutes, the VL3 loses all ethyl-lactate ligands. Then, slow condensation starts resulting in poly-vinyl-siloxan on the one hand. On the other hand, the formed ethyl-lactate slowly decomposes to ethanol and lactic acid. The rate constants are found in average 0.05 for the faster step and 0.002 min-1 for the slower one. The slower one is the base catalyzed decomposition of ethyl-lactate. At this pH VL3 hydrolysed first to ethyl lactate, then as result of ethyl-lactate hydrolysis, ethanol and lactic acid were found in the system. The concentrations at each time points are summarized in Table 10 while the actual amounts expressed as % (n/n) and % (m/m) of the original VL3 amount in Table 11 and 12. The signals of ethyl-lactate are so much overlapped that no quantititavie assessment is possible.

The first proposed hydrolysis step is the elimination of an ethyl-lactate ligand. From this point multiple parallel reaction routes are possible. The “opened” bond in the silicone centre is very reactive therefore it forms Si-O-Si bonds. If multiple ligands leave it results in the appearance of oligomeric and polymeric reaction products. The ethyl-lactate appearing in the spectra can also be the part of the chain when the polymerization starts and gradually gets free from the partially hydrolyzed oligomers. That is why the formation of ethyl-lactate can be detected at different rates. The signals originated from the colloid form appear as small intensity broad peaks at chemical shifts different from the non-polymerized VL3 or free ethyl-lactate. Further broadening effect is that the ester bond can be labile in the oligo- or polysiloxan and an exchange connection to the free ethyl-lactate results in wider signals. Parallel to this, the ethyl lactate might react further with water which usually leads to the formation of ethanol and lactic acid. The lactic acid might also form dilactide molcules. Further routes open when the remaining ethyl-lactate arms leave the molecule. The system behaviour was slightly different at the different pH values. The above described oligomer and polymer chains precipitate, therefore it was typical at all pH values that the overall NMR activity decreases, but with the dissolution of the immobilized ethyl-lactate functions the signal intensity increased over time.

Table 1: Hydrolysis product concentrations at each time points at pH 1.2:

pH=1.2 (mg/ml)

time (min)

ethanol

ethyl lactate

lactic acid

7

0.0

2.52

0.12

39

0.2

2.70

0.39

91

0.4

2.63

0.84

142

0.6

2.38

1.15

292

0.9

1.49

1.90

442

1.1

0.95

2.30

616

1.2

0.60

2.53

848

1.4

0.41

2.73

1021

1.4

0.35

2.80

1293

1.5

0.43

2.80

Table 2: Hydrolysis product actual amounts expressed as % (n/n) of the original VL3 amount at each time points at pH 1.2:

pH=1.2 (% n/n)

time (min)

ethanol

ethyl lactate

lactic acid

7

0.0

148.3

9.3

39

30.2

158.9

30.1

91

60.4

154.8

64.8

142

90.5

140.0

88.7

292

135.8

87.7

146.6

442

166.0

55.9

177.5

616

181.1

35.3

195.2

848

211.2

24.1

210.7

1021

211.2

20.6

216.1

1293

226.3

25.3

216.1

Table 3: Hydrolysis product actual amounts expressed as % (m/m) of the original VL3 amount at each time points at pH 1.2:

pH=1.2 (% m/m)

time (min)

ethanol

ethyl lactate

lactic acid

7

0.0

43.2

2.1

39

3.4

46.3

6.7

91

6.9

45.1

14.4

142

10.3

40.8

19.7

292

15.4

25.5

32.6

442

18.9

16.3

39.4

616

20.6

10.3

43.4

848

24.0

7.0

46.8

1021

24.0

6.0

48.0

1293

25.7

7.4

48.0

Table 4: Hydrolysis product concentrations at each time points at pH 4.0:

pH=4 (mg/ml)

time (min)

VL3

ethyl lactate

7

6.6

0.65

39

4.7

0.78

91

4.1

1.07

142

3.9

1.38

292

3.8

1.77

442

3.5

2.52

616

3.1

2.92

848

2.4

3.45

1021

1.5

4.67

1293

0.0

0.00

Table 5: Hydrolysis product actual amounts expressed as % (n/n) of the original VL3 amount at each time points at pH 4.0:

pH=4 (% n/n)

time (min)

VL3

ethyl lactate

7

84.3

28.5

39

60.0

34.2

91

52.3

46.9

142

49.8

60.5

292

48.5

77.6

442

44.7

110.4

616

39.6

127.9

848

30.6

151.2

1021

19.1

204.6

1293

0.0

0.0

Table 6: Hydrolysis product actual amounts expressed as % (m/m) of the original VL3 amount at each time points at pH 4.0:

pH=4 (% m/m)

time (min)

VL3

ethyl lactate

7

84.3

8.3

39

60.0

10.0

91

52.3

13.7

142

49.8

17.6

292

48.5

22.6

442

44.7

32.2

616

39.6

37.3

848

30.6

44.0

1021

19.1

59.6

1293

0.0

0.0

Table 7: Hydrolysis product concentrations at each time points at pH 7.0:

pH=7 (mg/ml)

time (min)

VL 3

ethyl lactate

3

4.1

1.5

20

2.3

1.9

51

1.0

2.6

72

0.4

2.6

132

0.0

3.0

222

0.0

3.2

372

0.0

3.5

672

0.0

4.0

972

0.0

4.4

Table 8: Hydrolysis product actual amounts expressed as % (n/n) of the original VL3 amount at each time points at pH 7.0:

pH=7 (% n/n)

time (min)

VL 3

ethyl lactate

3

60.0

75.3

20

33.7

95.4

51

14.6

130.6

72

5.9

130.6

132

0.0

150.7

222

0.0

160.7

372

0.0

175.8

672

0.0

200.9

972,00

0.0

221.0

Table 9: Hydrolysis product actual amounts expressed as % (m/m) of the original VL3 amount at each time points at pH 7.0:

pH=7 (% m/m)

time (min)

VL 3

ethyl lactate

3

60.0

22.0

20

33.7

27.8

51

14.6

38.0

72

5.9

38.0

132

0.0

43.9

222

0.0

46.8

372

0.0

51.2

672

0.0

58.5

972,00

0.0

64.4

Table 10: Hydrolysis product concentrations at each time points at pH 9.0:

pH=9 (mg/ml)

time (min)

VL 3

ethyl lactate

ethanol

3

4.3

1.24

0.00

10

2.2

1.71

0.00

18

1.5

1.80

0.00

33

0.0

2.12

0.04

44

0.0

2.04

0.04

144

0.0

2.09

0.13

287

0.0

1.98

0.21

576

0.0

1.69

0.36

986

0.0

1.50

0.47

Table 11: Hydrolysis product actual amounts expressed as % (n/n) of the original VL3 amount at each time points at pH 9.0:

pH=9 (% n/n)

time (min)

VL 3

ethyl lactate

ethanol

3

86,0

85,1

0,0

10

44,0

117,4

0,0

18

30,0

123,6

0,0

33

0,0

145,5

7,0

44

0,0

140,0

7,0

144

0,0

143,5

22,9

287

0,0

135,9

37,0

576

0,0

116,0

63,4

986

0,0

103,0

82,7

Table 12: Hydrolysis product actual amounts expressed as % (n/n) of the original VL3 amount at each time points at pH 9.0:

pH=9 (% m/m)

time (min)

VL 3

ethyl lactate

ethanol

3

86.0

24.8

0.0

10

44.0

34.2

0.0

18

30.0

36.0

0.0

33

0.0

42.4

0.8

44

0.0

40.8

0.8

144

0.0

41.8

2.6

287

0.0

39.6

4.2

576

0.0

33.8

7.2

986

0.0

30.0

9.4

Validity criteria fulfilled:
not specified
Conclusions:
The test item hydrolyses very fast at pH 1.2 (37 ºC), 4 (25 ºC), 7 (25 ºC) and 9 (25 ºC) being the half-lives 3.46, 4.62, 19.8 and 9.5 min respectively. The main hydrolysis product is ethyl lactate, but at pH 1.2 and 9.0 ethanol and lactic acid also appear. No other products were identified above 10% of the VL3 amount at the end of the hydrolysis. The presence of silane-dimer, trimer oligomer chains were seen in the beginning of the reactions. Since the NMR technique used is sensitive only for dissolved substances no quantitative information could be provided about these longer chain products, because they left the solution in form of colloids or gel-like white precipitate.
Executive summary:

An hydrolysis test was performed according to the OECD Guideline 111. Approximately 0.01 M solutions of the test item were prepared in buffers deoxygenated (by bubbling nitrogen through the buffers). The samples were kept isolated from light an oxygen in a drying oven at 25 ± 0.5°C (except at pH 1.2 which will be kept at 37°C) with daily temperature registration. Due to the complexity of the overall hydrolysis process it was hard to determine the reaction time necessary for the best description. The reactions were followed for approximately 24 hours. 1H NMR measurements were run consecutively using the same duplicate samples on a Bruker NMR spectrometer equipped with a z-gradient probehead. The first proposed hydrolysis step is the elimination of an ethyl-lactate arm. From this point multiple parallel reaction routes are possible. The “opened” bond in the silicone centre is very reactive therefore it forms various Si-O-Si bonds, which result in the appearance of polymeric reaction products. Parallel to this, ethyl lactate might react further with water which usually leads to the formation of ethanol and lactic acid. The lactic acid might also form dilactide molcules.

The system behaviour was slightly different at the different pH. In acidic medium (pH 1.2) the hydrolysis is fast and the ethyl-lactate has been released mostly (75% by 7 min). After this fast reaction step, the slow hydrolysis of ethyl-lactate to lactic acid and ethanol occurs. The half-life at pH 1.2 and 37 ºC (gastric conditions) was calculated to be of 3.46 min (k = 0.2 min-1). At pH 4, after the addition of the test item into the buffer, VL3 releases the ethyl-lactate and starts to condensate to vinyl-ethyl lactate colloid resulting in a decrease of VL3 concentration. Then the insoluble VL3 product undergoes further hydrolysis resulting in partial dissolution back to the solution, but in the same time it starts to hydrolyse and releases further free ethyl-lactate to the solution. At this pH VL3 hydrolysed only to ethyl-lactate. No ethanol or lactic acid was found in quantity higher than 1%. The half-life at pH 4 and 25 ºC was calculated to be of 4.62 min (k = 0.15 min-1). At pH 7, the first hydrolysis step is fast and results in consecutive loss of the ethyl-lactate arms from VL3 and in formation of oligomers. The rest of ethyl lactate probably dissolves out of the poly-vinyl-siloxan in a slower step during the condensation. At this pH VL3 hydrolysed only to ethyl lactate. No ethanol or lactic acid was found in quantity higher than 1%. The half-life at pH 7 and 25 ºC was calculated to be of 19.8 min (k = 0.035 min-1). The hydrolysis of VL3 in alcalic medium (pH 9) takes place in three steps. First quickly, within 40 minutes, the VL3 loses all ethyl-lactate ligands. Then slow condensation starts resulting in poly-vinyl-siloxanes on the one hand. On the other hand, the formed ethyl-lactate slowly decomposes to ethanol and lactic acid. Ethanol was found in the system, but interestingly no lactic acid was found in quantity higher than 1%. The half-life at pH 9 and 25 ºC was calculated to be of 9.5 min (k = 0.073 min-1).

Description of key information

Key study: OECD Guideline 111. The test item hydrolyses very fast at pH 1.2 (37 ºC), 4 (25 ºC), 7 (25 ºC) and 9 (25 ºC) being the half-lives 3.46, 4.62, 19.8 and 9.5 min respectively. The main hydrolysis product is ethyl lactate, but at pH 1.2 and 9.0 ethanol and lactic acid also appear. No other products were identified above 10% of the VL3 amount at the end of the hydrolysis. The presence of silane-dimer, trimer oligomer chains were seen in the beginning of the reactions. Since the NMR technique used is sensitive only for dissolved substances no quantitative information could be provided about these longer chain products, because they left the solution in form of colloids or gel-like white precipitate.

Key value for chemical safety assessment

Half-life for hydrolysis:
19.8 min
at the temperature of:
25 °C

Additional information

The key value for chemical safety assessment is determined to be 19.8 min (pH 9, 25 ºC) as the worst case (longest half-life from tested pHs).